Papers by Stuart Brierley
Brain, Behavior, and Immunity, 2017
Objective: Little is understood regarding how disease progression alters immune and sensory nerve... more Objective: Little is understood regarding how disease progression alters immune and sensory nerve function in colitis. We investigated how acute colitis chronically alters immune recruitment and the impact this has on reactivated colitis. To understand the impact of disease progress on sensory systems we investigated the mechanisms underlying altered colonic neuro-immune interactions after acute colitis. Design: Inflammation was compared in mouse models of Health, Acute trinitrobenzenesulphonic acid (TNBS) colitis, Remission and Reactivated colitis. Cytokine concentrations were compared by ELISA in-situ and in explanted colon tissue. Colonic infiltration by CD11b/F4-80 macrophage, CD4 T HELPER (T H) and CD8 T CYTOTOXIC (T C) and α 4 β 7 expression on mesenteric lymph node (MLN) T H and T C was determined by flow cytometry. Cytokine and effector receptor mRNA expression was determined on colo-rectal afferent neurons and the mechanisms underlying cytokinergic effects on high-threshold colo-rectal afferent function were investigated using electrophysiology. Results: Colonic damage, MPO activity, macrophage infiltration, IL-1β and IL-6 concentrations were lower in Reactivated compared to Acute colitis. T H infiltration and α 4 β 7 expression on T H MLN was increased in Remission but not Acute colitis. IFN-γ concentrations, T H infiltration and α 4 β 7 expression on T H and T C MLN increased in Reactivated compared to Acute colitis. Reactivated explants secreted more IL-1β and IL-6 than Acute explants. IL-6 and TNF-α inhibited colo-rectal afferent mechanosensitivity in Remission mice via a BK Ca dependent mechanism. Conclusions: Acute colitis persistently alters immune responses and afferent nerve signalling pathways to successive episodes of colitis. These findings highlight the complexity of viscero-sensory neuro-immune interactions in painful remitting and relapsing diseases.
The Open Pain Journal, 2013
Visceral pain is both different and similar to somatic pain - different in being poorly localized... more Visceral pain is both different and similar to somatic pain - different in being poorly localized and usually referred elsewhere to the body wall, but similar in many of the molecular mechanisms it employs (like TRP channels) and the specialization of afferent endings to detect painful stimuli. TRPV1 is sensitive to low pH. pH is lowest in gastric juice, which may cause severe pain when exposed to the oesophageal mucosa, and probably works via TRPV1. TRPV1 is found in afferent fibres throughout the viscera, and the TRPV1 agonist capsaicin can recapitulate symptoms experienced in disease. TRPV1 is also involved in normal mechanosensory function in the gut. Roles for TRPV4 and TRPA1 have also been described in visceral afferents, and TRPV4 is highly enriched in them, where it plays a major role in both mechanonociception and chemonociception. It may provide a visceral-specific nociceptor target for drug development. TRPA1 is also involved in mechano-and chemosensory function, but not ...
Gastroenterology
Increasing evidence suggests that chronic stress alters behavior and modifies epigenetic regulati... more Increasing evidence suggests that chronic stress alters behavior and modifies epigenetic regulation of genes in the central nervous system. DNA methylation, catalyzed by DNA methyltransferases (DNMTs), is an important epigenetic mechanism of transcriptional control of gene expression. We reported previously that chronic psychological stress induced visceral hyperalgesia and differential alterations in the expression of several genes in DRG neurons innervating the colon in the rat. A potential role for epigenetic regulation in peripheral sensory pathways has not been investigated. Objectives: We examined the hypothesis that DNMTs play an important role in the regulation of chronic stress-induced visceral hyperalgesia. Methods: Male rats were exposed to 1-hour water avoidance (WA) stress daily for 10 consecutive days as a chronic stress paradigm. SiRNA for DNMT1 was administrated in situ to L6-S2 DRGs every other day during the stress procedure. The visceromotor response (VMR) to colorectal distension was measured. Retrograde labeling with cholera toxin B (CTB)-FITC was employed to identify colon DRG neurons. Immunofluorescence and Western blot analysis were used to assess protein expression. In Vitro studies were performed in isolated control DRGs in the presence or absence of corticosterone (CORT; 10 μM) and RU-486 (corticoid receptor antagonist, 500 nM). Results: WA stress rats demonstrated significant increases in the level of DNMT1 and DNMT3b but not DNMT3a in L6-S2 DRGs compared with the controls. Enzyme activity assessment showed a 42% increase in DNMT1 activity in L6-S2 DRGs in stressed rats. Immunofluorescence studies revealed a significant increase in DNMT1 in small-sized, C-fiber neurons in WA stressed rats (52.3±2.2%) compared with the control (31.7±1.6%). Retrograde labeling demonstrated that 72.0±2.1% of the CTB-FITC labeled colonic DRG neurons were positive for DNMT1 in stressed rats compared to 40.0±6.5% in controls (P<0.05; n=4). The VMR in WA stressed rats was increased 68% and 92% above control responses at pressures of 40 and 60 mm Hg, respectively. Treatment of stressed rats with siRNA for DNMT1 prevented the VMR enhancement and changes in DNMT1 proteins levels in L6-S2 DRGs. In addition, treatment of control L6-S2 DRGs In Vitro with CORT (10 μM) increased DNMT1 expression level that was prevented by RU-486 (500 nM) (P<0.05). Conclusions: These data support the novel and provocative interpretation that: 1. Chronic stress induces epigenetic regulation of genes in primary nociceptive neurons; 2. DNA methyltransferase 1 (DNMT1) plays an important role in modulation of chronic stress-induced visceral hyperalgesia; and 3. DNA methyltransferases represent a potential target for treatment of functional GI disorders associated with visceral hyperalgesia.
Chronic abdominal pain or discomfort is common and frequently these patients seek health care (1)... more Chronic abdominal pain or discomfort is common and frequently these patients seek health care (1). In >50 % of patients presenting with chronic abdominal symptoms, no structural lesion is found aft er a standard diagnostic work-up that is likely to explain these symptoms (2). Consequently, patients are diagnosed as having a functional gastrointestinal disorder (FGID); the best-recognized disorders are the irritable bowel syndrome (IBS) and functional dyspepsia (FD) (3). Unfortunately, the underlying pathophysiological mechanisms in IBS and FD still remain poorly understood.
Brain, Behavior, and Immunity, 2014
Alterations in the neuro-immune axis contribute towards viscerosensory nerve sensitivity and symp... more Alterations in the neuro-immune axis contribute towards viscerosensory nerve sensitivity and symptoms in Irritable Bowel Syndrome (IBS). Inhibitory factors secreted from immune cells inhibit colo-rectal afferents in health, and loss of this inhibition may lead to hypersensitivity and symptoms. We aimed to determine the immune cell type(s) responsible for opioid secretion in humans and whether this is altered in patients with IBS. The β-endorphin content of specific immune cell lineages in peripheral blood and colonic mucosal biopsies were compared between healthy subjects (HS) and IBS patients. Peripheral blood mononuclear cell (PBMC) supernatants from HS and IBS patients were applied to colo-rectal sensory afferent endings in mice with post-inflammatory chronic visceral hypersensitivity (CVH). βendorphin was identified predominantly in monocyte / macrophages relative to T or B cells in human PBMC and colonic lamina propria. Monocyte derived β-endorphin levels and colonic macrophage numbers were lower in IBS patients than healthy subjects. PBMC supernatants from healthy subjects had greater inhibitory effects on colo-rectal afferent mechanosensitivity than those from IBS patients. The inhibitory effects of PBMC supernatants were more prominent in CVH mice compared to healthy mice due to an increase in µ-opioid receptor expression in dorsal root ganglia neurons in CVH mice. Monocyte / macrophages are the predominant immune cell type responsible for β-endorphin secretion in humans. IBS patients have lower monocyte derived β-endorphin levels than healthy subjects, causing less inhibition of colonic afferent endings. Consequently, altered immune function contributes toward visceral hypersensitivity in IBS.
Zeitschrift für Gastroenterologie, 2007
The Journal of Physiology, 2011
Non-technical summary Detecting mechanical stimuli is vital to determining our responses to envir... more Non-technical summary Detecting mechanical stimuli is vital to determining our responses to environmental challenges. The speed required for this process suggests ion channels are opened in response to mechanical forces. A specialised membrane protein called the TRPA1 ion channel mediates chemical based pain; however its role in mechanical pain remains unresolved. Here we show that TRPA1 contributes to the detection of mechanical stimuli in a select set of pain sensing neurons. Furthermore, we also show that acute activation of this channel enhances the mechanical responsiveness of these neurons. Finally, we also show that increasing the expression of TRPA1 causes a further enhancement in the mechanical response. These findings suggest that a drug designed to block TRPA1 would be beneficial for the treatment of numerous pathological conditions associated with mechanical pain.
The Journal of Physiology, 2006
Glutamate acts at central synapses via ionotropic (iGluR-NMDA, AMPA and kainate) and metabotropic... more Glutamate acts at central synapses via ionotropic (iGluR-NMDA, AMPA and kainate) and metabotropic glutamate receptors (mGluRs). Group I mGluRs are excitatory whilst group II and III are inhibitory. Inhibitory mGluRs also modulate peripherally the mechanosensitivity of gastro-oesophageal vagal afferents. Here we determined the potential of excitatory GluRs to play an opposing role in modulating vagal afferent mechanosensitivity, and investigated expression of receptor subunit mRNA within the nodose ganglion. The responses of mouse gastro-oesophageal vagal afferents to graded mechanical stimuli were investigated before and during application of selective GluR ligands to their peripheral endings. Two types of vagal afferents were tested: tension receptors, which respond to circumferential tension, and mucosal receptors, which respond only to mucosal stroking. The selective iGluR agonists NMDA and AMPA concentration-dependently potentiated afferent responses. Their corresponding antagonists AP-5 and NBQX alone attenuated mechanosensory responses as did the non-selective antagonist kynurenate. The kainate selective agonist SYM-2081 had minor effects on mechanosensitivity, and the antagonist UBP 302 was ineffective. The mGluR5 antagonist MTEP concentration-dependently inhibited mechanosensitivity. Efficacy of agonists and antagonists differed on mucosal and tension receptors. We conclude that excitatory modulation of afferent mechanosensitivity occurs mainly via NMDA, AMPA and mGlu5 receptors, and the role of each differs according to afferent subtypes. PCR data indicated that all NMDA, kainate and AMPA receptor subunits plus mGluR5 are expressed, and are therefore candidates for the neuromodulation we observed.
The Journal of Physiology, 2013
• Obesity occurs when energy intake exceeds expenditure, and the excess energy is stored as fat. ... more • Obesity occurs when energy intake exceeds expenditure, and the excess energy is stored as fat. • We show that, after a 14 h food deprivation or 12 weeks consumption of a high-fat diet, gastric vagal afferent responses to mechanical stimulation in the presence of the satiety peptide leptin are altered. • Leptin has an excitatory effect on gastric mucosal vagal afferents, which is abolished after food restriction or prolonged excess. • In contrast, leptin has an inhibitory effect on gastric tension-sensitive afferents, but only after food restriction or energy excess conditions. • These changes in the response to leptin in the stomach, after food restriction or prolonged high-fat feeding, occur in such a manner as to facilitate an increase in food intake in both conditions.
The Journal of Physiology, 2007
Pharmaceuticals, 2010
We are normally unaware of the complex signalling events which continuously occur within our inte... more We are normally unaware of the complex signalling events which continuously occur within our internal organs. Most of us only become cognisant when sensations of hunger, fullness, urgency or gas arise. However, for patients with organic and functional bowel disorders pain is an unpleasant and often debilitating reminder. Furthermore, chronic pain still represents a large unmet need for clinical treatment. Consequently, chronic pain has a considerable economic impact on health care systems and the afflicted individuals. In order to address this need we must understand how symptoms are generated within the gut, the molecular pathways responsible for generating these signals and how this process changes in disease states.
Pain, 2008
Metabotropic glutamate 5 receptor (mGluR5) antagonists are effective in animal models of inflamma... more Metabotropic glutamate 5 receptor (mGluR5) antagonists are effective in animal models of inflammatory and neuropathic pain. The involvement of mGluR5 in visceral pain pathways from the gastrointestinal tract is as yet unknown. We evaluated effects of mGluR5 antagonists on the colorectal distension (CRD)-evoked visceromotor (VMR) and cardiovascular responses in conscious rats, and on mechanosensory responses of mouse colorectal afferents in vitro. Sprague-Dawley rats were subjected to repeated, isobaric CRD (12 • 80 mmHg, for 30 s with 5 min intervals). The VMR and cardiovascular responses to CRD were monitored. The mGluR5 antagonists MPEP (1-10 lmol/kg, i.v.) and MTEP (1-3 lmol/kg, i.v.) reduced the VMR to CRD dose-dependently with maximal inhibition of 52 ± 8% (p < 0.01) and 25 ± 11% (p < 0.05), respectively, without affecting colonic compliance. MPEP (10 lmol/kg, i.v.) reduced CRD-evoked increases in blood pressure and heart rate by 33 ± 9% (p < 0.01) and 35 ± 8% (p < 0.05), respectively. Single afferent recordings were made from mouse pelvic and splanchnic nerves of colorectal mechanoreceptors. Circumferential stretch (0-5 g force) elicited slowly-adapting excitation of action potentials in pelvic distension-sensitive afferents. This response was reduced 55-78% by 10 lM MTEP (p < 0.05). Colonic probing (2 g von Frey hair) activated serosal splanchnic afferents; their responses were reduced 50% by 10 lM MTEP (p < 0.01). We conclude that mGluR5 antagonists inhibit CRD-evoked VMR and cardiovascular changes in conscious rats, through an effect, at least in part, at peripheral afferent endings. Thus, mGluR5 participates in mediating mechanically evoked visceral nociception in the gastrointestinal tract.
Pain, 2011
Transient receptor potential ion channel melastatin subtype 8 (TRPM8) is activated by cold temper... more Transient receptor potential ion channel melastatin subtype 8 (TRPM8) is activated by cold temperatures and cooling agents, such as menthol and icilin. Compounds containing peppermint are reported to reduce symptoms of bowel hypersensitivity; however, the underlying mechanisms of action are unclear. Here we determined the role of TRPM8 in colonic sensory pathways. Laser capture microdissection, quantitative reverse transcription-polymerase chain reaction (RT-PCR), immunofluorescence, and retrograde tracing were used to localise TRPM8 to colonic primary afferent neurons. In vitro extracellular single-fibre afferent recordings were used to determine the effect of TRPM8 channel activation on the chemosensory and mechanosensory function of colonic high-threshold afferent fibres. TRPM8 mRNA was present in colonic DRG neurons, whereas TRPM8 protein was present on nerve fibres throughout the wall of the colon. A subpopulation (24%, n = 58) of splanchnic serosal and mesenteric afferents tested responded directly to icilin (5 lmol/L). Subsequently, icilin significantly desensitised afferents to mechanical stimulation (P < .0001; n = 37). Of the splanchnic afferents responding to icilin, 21 (33%) also responded directly to the TRPV1 agonist capsaicin (3 lmol/L), and icilin reduced the direct chemosensory response to capsaicin. Icilin also prevented mechanosensory desensitization and sensitization induced by capsaicin and the TRPA1 agonist AITC (40 lmol/L), respectively. TRPM8 is present on a select population of colonic high threshold sensory neurons, which may also co-express TRPV1. TRPM8 couples to TRPV1 and TRPA1 to inhibit their downstream chemosensory and mechanosensory actions.
Neurogastroenterology and Motility, 2005
Lumbar splanchnic (LSN) and sacral pelvic (PN) nerves convey different mechanosensory information... more Lumbar splanchnic (LSN) and sacral pelvic (PN) nerves convey different mechanosensory information from the colon to the spinal cord. Here, we determined whether these pathways differ also in their chemosensitivity to bradykinin. Using a novel in vitro mouse colon preparation, serosal afferents were recorded from the LSN and PN and distinguished based on their mechanosensitivity to von Frey filaments (70-4000 mg) and insensitivity to colonic stretch (1-5 g) or fine mucosal stroking (10 mg). Bradykinin was applied into a ring around mechanoreceptive fields. The LSN and PN afferents had different dynamic responses to mechanical stimuli: PN afferents required lower intensity stimuli, evoked larger responses, and displayed more maintained responses than LSN afferents. Bradykinin (1 micromol L-1) excited 66% (27 of 41) of LSN afferents. Responses to probing were potentiated after bradykinin. The concentration-dependent (EC50: 0.16 micromol L-1) response was reversed by the B2-receptor antagonist HOE-140 (10 nmol L-)). Twelve bradykinin responsive afferents were mechanically insensitive. More LSN serosal afferents responded to bradykinin than PN afferents (11%, P&lt;0.001) , with larger responses (P&lt;0.05). No mechanically insensitive PN afferents were recruited by bradykinin. Bradykinin potently stimulates most splanchnic serosal afferents via B2-receptors, but few pelvic afferents. Mechanically insensitive afferents recruited by bradykinin are exclusive to the LSN.
Neurogastroenterology & Motility, 2013
Background: The transient receptor potential vanilloid 1 (TRPV1) channel is critical for spinal a... more Background: The transient receptor potential vanilloid 1 (TRPV1) channel is critical for spinal afferent signalling of burning pain throughout the body. Such pain frequently originates from the esophagus, following acid reflux. The contribution of TRPV1 to spinal nociceptor signalling from the esophagus remains unclear. We aimed to identify the spinal afferent pathways that convey nociceptive signalling from the esophagus, specifically those sensitive to acid, and the extent to which TRPV1 contributes. Methods: Acid/pepsin (150mM HCl / 1mg ml-1 pepsin) or saline/pepsin was perfused into the esophageal lumen of anesthetised wild-type and TRPV1 null mice over 20 minutes, followed by atraumatic perfuse-fixation and removal of the cervical and thoracic spinal cord and dorsal root ganglia (DRG). To identify neurons responsive to esophageal perfusate, immunolabelling for neuronal activation marker phosphorylated extracellular receptorregulated kinase (pERK) was used. Labelling for calcitonin gene related peptide (CGRP) and isolectin B4 (IB4) was then used to characterize responsive neurons. Key Results: Esophageal acid/pepsin perfusion significantly increased the number of pERKimmunoreactive (IR) neurons in the DRG and the cervical and thoracic spinal cord dorsal horn (DH) relative to saline/pepsin (DRG P<0.01; cervical DH P<0.05 and thoracic DH P<0.005). The number of pERK-IR neurons following acid perfusion was significantly attenuated in TRPV1-/-mice (DH P<0.05 and DRG P<0.05). Conclusions and inferences: This study has identified populations of spinal afferent DRG neurons and DH neurons involved in signalling of noxious acid from the esophagus. There is a major contribution of TRPV1 to signalling within these pathways.
Gut, 2009
The paper by Akbar et al (Gut 2008;57:9239) characterises the expression of the capsaicin recept... more The paper by Akbar et al (Gut 2008;57:9239) characterises the expression of the capsaicin receptor, transient receptor potential vanilloid type-1 (TRPV1), immune cell markers, and their relationship to symptoms in irritable bowel syndrome (IBS). They conclude that the ...
Gastroenterology, 2008
which increased to 10-12% of all DCs. However, the increase in CD103+DCs was shortlived. Instead,... more which increased to 10-12% of all DCs. However, the increase in CD103+DCs was shortlived. Instead, the healing phase of colitis was characterized by the influx of pDCs which increased over 10-fold during a 7 day recovery period and stayed elevated for over 3 weeks. The influx of pDCs was accompanied by the recruitment of Foxp3+Tregs in the colon which were found dispersed throughout the lamina propria and steadily increased during mucosal healing. Surprisingly, IL-10 deficient mice demonstrated the enhanced recruitment of pDCs (25-30% of all DCs) and Foxp3+Tregs even before the onset of colitis but without an increased in CD103+DCs. Onset of macroscopic disease in IL-10 deficient mice was characterized by the additional influx of CD11b-DCs. CONCLUSION: Mucosal defense activation involves the recruitment of specific DC subsets which have defined roles in the control of acute inflammation and support of mucosal healing. Surprisingly, DC subset recruitment was similar in colitis induced by IEC monolayer disruption and T cell mediated disease in IL-10 deficient mice. This suggests common response programs to mucosal damage. We would like to propose a model in which a distinct CD103+DC subset in ILFs is responsible for the induction of Foxp3+Tregs but pDCs are required for the expansion and maintenance of Tregs in mucosal healing.
Gastroenterology, 2008
Background: There is increasing evidence that inflammation and immune activation may contribute t... more Background: There is increasing evidence that inflammation and immune activation may contribute to the manifestation of symptoms in at least a subgroup of patients with Functional Dyspepsia (FD). However, very little is known about cellular immune activation in FD and its association with symptom manifestation. Detection of alpha-4 integrin (α4) and beta-7 integrin (β7) on circulating T cells identifies specific lymphocyte subsets trafficking between the circulation and the gut. In addition CCR9 expression discriminates between small and large "bowel homing" lymphocytes. We hypothesized that patients with FD have increased circulating 'gut homing' T cells which might be linked to symptom severity. Thus we aimed to quantitate a) circulating CD4+T cells numbers, b) α4+β7+ positive "large bowel homing" T cells and c) α4+β7+CCR9+ "small bowel homing" T cell subsets in patients with FD compared to healthy controls and to correlate gut homing T cells with symptom severity. Methods: Peripheral blood mononuclear cells (PBMC) from 28 (H. pylori negative) FD (ROME II) patients and 26 matched healthy controls (HC) were isolated by density gradient centrifugation. Cell surface expression of CD4+ T cells, alpha-4 integrin, beta-7 integrin and CCR9 was analyzed by four color flow cytometry, while quantitative α-4 integrin, β-7integrin and CCR9 mRNA expression were characterized by RT-PCR in comparison to 4 different plasmid standards. Abdominal symptoms were characterized by a standardized Gastrointestinal Symptom Score. Results: Overall no significant differences were observed for numbers of CD4+ T cells and CD4+ α4+β7+ (50.3±6.4 vs. 42.8±5.9%) lymphocytes between patients and HC. Small bowel homing CD4+ α4+β7+CCR9+ T lymphocytes were significantly (p<0.05) increased in patients with in FD (5.6±1.4 vs. 1.6±0.4%). CCR9, but not α4 or β7 integrin mRNA expression was significantly higher in FD compared to HC. However, no correlation was observed between CD4+ α4+β7+CCR9+ lymphocytes and symptom severity. Summary and Conclusion: Patients with FD are characterized by increased circulating small bowel homing T lymphocytes. These findings are consistent with an underlying immune activation in FD. The role in the manifestation of symptoms needs to be further elucidated.
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Papers by Stuart Brierley